tbuckley/cabot-search huggingface dataset:
Abstract P3199: Role Of Acetylation In Regulating Mitochondrial Dysfunction In The Aged Heart
Authors: Jackson E Stewart, Jenna M Crawford, Angelica A Jacques, Michael W Stoner, Iain Scott, Dharendra Thapa
Journal: Circulation Research (2023)

Cardiac mitochondrial dysfunction is a critical contributor to the pathogenesis of aging. Cardiac muscle is an extremely high energy demanding tissue, necessitating properly functioning mitochondria to meet this demand for contractile function. As such, complete control of mitochondrial function is indispensable to maintain cardiac efficiency. Lysine acetylation is a post-translational modification of proteins that has been shown to regulate several metabolic and biochemical processes in the mitochondria. The goal of this study is to understand the role of mitochondrial lysine acetylation in regulating mitochondrial processes in the aged heart. We observed an increased propensity towards a pro-acetylation profile resulting in increased acetylation of several cardiac mitochondrial proteins involved in metabolism, redox milieu and electron transport chain complexes in aging which was attenuated in our cardiac GCN5L1 knockout old animal. Based upon these findings, we hypothesize that mitochondrial acetyltransferase GCN5L1 has a critical role in regulating the function of mitochondrial proteins in the aged heart. Using immunoprecipitation assay, we observed an age-associated increase in acetylation of several key fatty acid oxidation and glucose oxidation proteins. This increased acetylation correlated with decreased activity of several fatty acid oxidation proteins, including short- and long-chain acyl-CoA dehydrogenase. Several mitochondrial proteins involved in mitochondrial fatty acid import and electron transport chain complexes were hyperacetylated in aging. These proteins largely exhibited increased acetylation levels in response to age that was attenuated by cardiac-specific GCN5L1 knockout. Lastly, we observed an age-associated increase in GCN5L1-mediated acetylation of the antioxidant enzyme manganese superoxide dismutase (SOD2) at lysine site K122. Acetylation of SOD2 at the K122 site has been shown to inhibit SOD2 activity. Based upon our findings, we conclude that age-associated increases in GCN5L1 expression results in derangements in cardiac fuel substrate utilization, electron transport chain complex function, and mediation of reactive oxygen species-associated damage.
Abstract EC.02: Myosin Light Chain Dephosphorylation By Ppp1r12c Promotes Atria Hypocontractility In Atrial Fibrillation
Authors: Francisco J Gonzalez-Gonzalez, srikanth perike, Issam Abu-Taha, Frederick Damen, Ken S Lizama, Anahita Aboonabi, Andrielle Capote, Yuriana Aguilar-Sanchez, Benjamin Levin, Zhenbo Han, Arvind Sridhar, Jacob Grand, Jody C Martin, Joseph Akar, Chad M Warren, R John Solaro, Sang-Ging Ong, Dawood Darbar, Craig J Goergen, Beata M Wolska, Dobromir Dobrev, Xander H Wehrens, Mark McCauley
Journal: Circulation Research (2023)

Background: Atrial fibrillation (AF), the most common sustained cardiac arrhythmia, increases thromboembolic stroke risk five-fold. Although atrial hypocontractility contributes to stroke risk in AF, the molecular mechanisms reducing myofilament contractile function remain unknown. We tested the hypothesis that increased expression of PPP1R12C, the PP1 regulatory subunit targeting atrial myosin light chain 2 (MLC2a), causes hypophosphorylation of MLC2a and results in atrial hypocontractility. Methods: Right atrial appendage tissues were isolated from human AF patients versus sinus rhythm (SR) controls. Western blots, co-immunoprecipitation, and phosphorylation studies were performed to examine how the PP1c-PPP1R12C interaction causes MLC2a de-phosphorylation. In vitro studies of pharmacologic MRCK inhibitor (BDP5290) in atrial HL-1 cells were performed to evaluate PP1 holoenzyme activity on MLC2a. Cardiac-specific lentiviral PPP1R12C overexpression was performed in mice to evaluate atrial remodeling with atrial cell shortening assays, echocardiography, and AF inducibility with EP studies. Results: In human patients with AF, PPP1R12C expression was increased two-fold versus SR controls with &gt; 40% reduction in MLC2a phosphorylation. 12C-PP1c binding and PPP1R12C-MLC2a binding were significantly increased in AF. In vitro studies utilizing drug BDP5290, which inhibits T560-PPP1R12C phosphorylation, demonstrated increased PPP1R12C binding with both PP1c and MLC2a, and dephosphorylation of MLC2a. Lenti-12C mice demonstrated a 150% increase in LA size versus controls, with reduced atrial strain and atrial ejection fraction. Pacing-induced AF in Lenti-12C mice was significantly higher than controls. Conclusions: AF patients exhibit increased levels of PPP1R12C protein compared to controls. PPP1R12C overexpression in mice increases PP1c targeting to MLC2a and causes MLC2a dephosphorylation, which reduces atrial contractility and increases AF inducibility. These findings suggest that PP1 regulation of sarcomere function at MLC2a is a key regulator of atrial contractility in AF.
Abstract P2059: Perm1 Regulates O-glcnacylation Of Pgc-1alpha In The Heart
Authors: KARTHI SREEDEVI, Shreya Yedla, James Amina, Sumaita Arowa, Riyan Shah, Sara Do, Breanna Goode, Sydney Bui, Katia Olmos, Alexey Zaitsev, Junco Warren
Journal: Circulation Research (2023)

O-GlcNAcylation (O-GlcNAc) is a post-translational modification of proteins where N-acetylglucosamines are added to serine and threonine residues in proteins. This modification is regulated by two enzymes: O-GlcNAc transferase (OGT) that adds O-GlcNAc to proteins; O-GlcNAcase (OGA) that removes O-GlcNAc from proteins. A recent study demonstrated that overexpression of OGT in the heart leads to excessive O-GlcNAcylation, resulting in mitochondrial dysfunction and the development of heart failure. However, what regulates OGT expression in the heart remains elusive. Perm1 is a striated-muscle-specific regulator of mitochondrial bioenergetics. Here, we found that Perm1 suppresses O-GlcNAcylation via gene repression of OGT. Adenovirus-mediated overexpression of Perm1 suppressed O-GlcNAcylation of proteins in H9c2 cells (47% of control, p&lt;0.05), concurrent with downregulation of OGT (52% from control, p&lt;0.05). Luciferase gene reporter assay revealed that Perm1 reduces the promoter activity of OGT, indicating that Perm1 acts as a gene repressor of OGT. Furthermore, phenylephrine-induced O-GlcNAcylation in cardiomyocytes was reversed by Perm1 overexpression through repressing OGT expression. Conversely, loss of Perm1 (Perm1-KO) in mice led to upregulation of OGT and the increase of O-GlcNAcylated proteins in the heart (176% and 144% from WT, respectively, both p&lt;0.05, n=6 in WT, n=7 in KO). Concomitantly, Perm1-KO mice exhibited reduced contractility and mitochondrial function, manifested by a significant decrease in ejection fraction and succinate dehydrogenase activity, respectively (87% and 53% of WT, both p&lt;0.05). Lastly, subcellular fractionation and co-immunoprecipitation assay of mouse hearts revealed that loss of Perm1 specifically increases O-GlcNAcylation of nuclear proteins, concurrent with excessive O-GlcNAcylation of PGC-1α, a master regulator of mitochondrial bioenergetics (2-fold increase in O-GlcNAcylated PGC-1α/total PGC-1α, p&lt;0.05, n=3/group). Our results suggest that Perm1 is a novel regulator of O-GlcNAcylation in the heart that transcriptionally represses OGT expression, which presumably prevents excessive O-GlcNAcylation of PGC-1α and mitochondrial dysfunction under pathological stress.
Abstract P3048: <i>Ppp1r1b-lncRNA</i> Regulates Cardiomyocytes Differentiation Via Epigenetic Modulation Of <i>Tbx5</i> Promoter
Authors: Xuedong Kang, John Hwang, Marlin Touma
Journal: Circulation Research (2023)

Introduction: Myogenic differentiation is essential to cardiac development as well as regeneration and repair in response to genetic lesions or injury in the context of cardiomyopathy and congenital heart defects (CHDs). We have previously shown that mouse Ppp1r1b-lncRNA modulates epigenetic features of myogenic master regulators through targeted interaction with histone-modifying complex and other co-factors. However, the molecular mechanisms that underlie human PPP1R1B-lncRNA -mediated regulation of cardiomyocyte differentiation remain unknown. Objectives: To elucidate the mechanisms by which human PPP1R1B-lncRNA regulates cardiomyocyte differentiation. Methods/Results: PPP1R1B-lncRNA possessed conserved function in promoting myogenic differentiation in human cardiac and skeletal myocytes and its expression was dysregulated in CHDs. Mechanistically, by silencing PPP1R1B-lncRNA , myogenic differentiation was impaired in PPP1R1B-lncRNA -deficient human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs). The expressions of myogenic transcription factors, including GATA4 and TBX5 , as well as sarcomere proteins, including TNNT2 and MYOZ2, were significantly suppressed in PPP1R1B-lncRNA deficient hiPSC-CMs. Epigenetic analysis revealed increased H3K27 tri-methylation at TBX5 promoter in the PPP1R1B-lncRNA knockdown cardiac progenitors. PPP1R1B-lncRNA was found to bind to EZH2, the enzymatic core of the polycomb repressive complex (PRC2). Furthermore, chromatin isolation by RNA purification (ChIRP)-seq revealed enriched interaction of PPP1R1B-lncRNA with TBX5 , MEF2C, and GATA4 promoters. Meanwhile, chromatin immunoprecipitation (ChIP)-PCR demonstrated increased enrichments of EZH2 at TBX5 promoter in the PPP1R1B-lncRNA-depleted cells, resembling what was observed in the pre-differentiation state. Conclusions: Human PPP1R1B-lncRNA promotes cardiomyocyte differentiation via interactions with promoters of key cardiac transcription factors. PPP1R1B-lncRNA enhances chromatin accessibility at TBX5 promoter by interfering with PRC2 binding. Understanding PPP1R1B-lncRNA function in cardiomyocyte differentiation may lead to novel therapies for cardiopathies and CHDs.
Abstract P2061: The Intersection Of O-GlcNAc And Yap - A Novel Molecular Axis In Heart Failure
Authors: Gabriel Lopez-Cecetaite, Elizabeth Luczak, Priya Umapathi
Journal: Circulation Research (2023)

Introduction: Heart failure (HF) and pathological myocardial hypertrophy continue to grow world-wide and are strongly tied to obesity, hyperglycemia, and ischemia. These stressors lead to increased levels of O-beta-N-Acetylglucosamine modified proteins (OGN) in the heart. OGN levels are determined by the net activity of two enzymes, OGT and OGA. Our recent work showed excessive OGN was sufficient to cause HF and pathological myocardial hypertrophy. Here we test the relationship between OGN and Yes Associated Protein (YAP), a known promoter of cardiac hypertrophy. Objectives: Test if OGN modifies YAP in the heart and evaluate the role of OGN modified YAP in the development of pathologic cardiac hypertrophy and heart failure. Methods: We have shown that mouse models with myocardial OGT overexpression (OGT TG) developed pathological cardiac hypertrophy and HF. We found these animals have increased YAP activity. Myocardial OGA overexpression (OGA TG) attenuated OGN following transaortic constriction(TAC) and were protected against cardiac hypertrophy and HF. Interbreeding OGT TG with OGA TG rescued animals from pathological hypertrophy, HF and normalized YAP activity . We quantified levels of activated YAP protein/gene expression in hearts from WT, OGT, OGA and OGT x OGA mice. We evaluated YAP/TEAD1 target gene expression levels in OGA TG mice protected against TAC. We used an adenoviral YAP/TEAD expression reporter to quantify YAP/TEAD1 activity in cardiac myocytes, in the presence and absence of increased O-GlcNAcylation. Results: We found WT levels of activated (nuclear) YAP in OGA TG, increased active YAP in OGT TG and normal levels in OGT x OGA mice. YAP/TEAD1 target genes are upregulated in OGT TG hearts but interbred OGT x OGA animals have normal cardiac function and YAP/TEAD1 target gene expression. OGA TG mice who are protected against TAC induced heart failure/hypertrophy and have decreased myocardial levels of YAP/TEAD1 gene targets involved in pathological hypertrophy. Increased O-GlcNAcylation in cardiac myocytes activates YAP/TEAD activity and YAP/TEAD downstream gene expression. Conclusion: These preliminary data suggest a novel pathway for promoting pathological hypertrophy and HF via activation of the O-GlcNAc -YAP/TEAD1 axis.
Abstract P2159: Naltrindole Attenuates Phorbol 12-myristate 13-acetate Induced Superoxide Release In Polymorphonuclear Leukocytes Independently Of Delta Opioid Receptor Antagonism
Authors: Devani Johnson, Taylor DiLisi, Juliet Melnik, Denise Pinto, Alexandra Barrera, Arjun Nair, Zinya Talukder, Tameka Dean, Qian CHEN, Robert Barsotti, Lindon Young
Journal: Circulation Research (2023)

Naltrindole (NTI) is a selective delta opioid receptor antagonist that recently has been shown to be cardioprotective in both in vivo and ex vivo studies. In vivo, NTI showed cardioprotective effects that were dose-dependent significantly reducing infarct size up to 82% compared to controls subjected to ischemia-reperfusion (I/R) injury (p&lt;0.05). In ex vivo isolated rat I/R hearts, NTI significantly reduced infarct size compared to controls (p&lt;0.01). NTI restored cardiac left ventricular end diastolic pressure to near baseline values compared to controls or other general opioid receptor antagonists in ex vivo hearts, suggesting a tissue salvaging mechanism independent of opioid delta receptor antagonism. Polymorphonuclear leukocytes (PMNs) do not express opioid receptors. In this study, we investigated NTI attenuation of PMN superoxide (SO) release in vitro . We hypothesized that NTI would attenuate PMN SO release via a calcium handling mechanism, a similar mechanism that explained cardioprotective effects of NTI pretreatment mitigating ischemic hypercontracture due to an increase in intracellular calcium. Rat PMNs (5x10 6 ) were incubated for 15 min at 37 o C in the presence or absence (dH 2 O vehicle controls) of NTI (10μM, 50μM, or 200μM). PMN SO release was calculated by the change in absorbance at 550 nm over 420 sec via ferricytochrome c reduction after phorbol 12-myristate-13 acetate (PMA) stimulation (100nM). Cell viability was determined microscopically by 0.2% trypan blue exclusion at the end of the assay. Data were analyzed using ANOVA Fisher’s PLSD post-hoc test. NTI significantly decreased PMN SO release from 30 sec to 420 sec. Changes in final absorbance of NTI 200μM (0.244±0.07, n=8, p&lt;0.05) and 50μM (0.293±0.06, n=4, p&lt;0.05), but not NTI 10μM (0.412±0.07, n=5), were significant compared to controls (0.487±0.12, n=9). Cell viability was not significantly different compared to PMA across all NTI concentrations. These results suggest that NTI reduces I/R injury and PMN SO release, by a means independent of delta opioid receptor antagonism. Future studies will assess NTI SO attenuation in human umbilical vein endothelial cells. We will use different concentrations to investigate optimal concentration response.
Abstract P3075: Neutrophil Secreted Chitinase 3-like-1 Antagonizes Macrophage Reparative Polarization And Contributes To Ventricular Dysfunction After Myocardial Infarction
Authors: Jonah Stephan, Taylor Zoubul, Yibing Nong, Robert Brainard, Marcin Wysoczynski
Journal: Circulation Research (2023)

Clinical studies show that chitinase 3-like-1 (CHI3L1) is a biomarker and predictor of all-cause mortality in heart failure. Studies from our lab demonstrated that CHI3L1 is elevated in hearts post-MI, and CHI3L1 deficiency in mice results in preserved ventricular structure and function post-MI. Furthermore, we found that CHI3L1 is secreted from activated neutrophils but has no effect on neutrophil recruitment in hearts after MI. The goal of the current study was to elucidate mechanisms whereby CHI3L1 contributes to exacerbated cardiac dysfunction after MI. Post-MI neutrophil recruitment is followed by the sequential programming of macrophages to pro-inflammatory or pro-reparative phenotypes. We sought to determine the impact of neutrophil-secreted CHI3L1 on macrophage polarization. Activated neutrophils isolated from Chil1 +/+ and Chil1 -/- mice were co-cultured with bone marrow-derived macrophages. We found that macrophages directly co-cultured with Chil1 -/- but not Chil1 +/+ neutrophils showed upregulation of the pro-reparative markers, Chil4 and Arg1 (n=5/group, p&lt;0.05). Additionally, direct co-culture of Chil1 -/- neutrophils and macrophages showed trends towards downregulation of the pro-inflammatory polarization markers, Il1b and Tnfa relative to macrophages co-cultured with Chil +/+ neutrophils. These data suggest that neutrophil-derived CHI3L1 may antagonize signaling to polarize macrophages towards a pro-reparative phenotype that may explain delayed scar formation and exacerbated ventricular dysfunction after MI. Future efforts will elucidate the signaling role of CHI3L1 associated with neutrophil extracellular traps (NETs) on macrophage function.
Abstract P3190: Renal Tubular Mitochondrial Akt1 Modulates Cardiorenal Metabolic Syndrome
Authors: Kai-Ting Chang, Hugo Y Lin
Journal: Circulation Research (2023)

Metabolic syndrome-induced cardiomyopathy is a significant risk factor for cardiovascular diseases (CVDs). The proximal tubular cells of the kidney play a crucial role in regulating glucose and lipid metabolism. Mitochondrial dysfunction in these cells has been linked to the development of metabolic syndrome-induced cardiac dysfunction. AKT1, a serine/threonine protein kinase, regulates mitochondrial function. In this study, we investigated the effects of renal proximal tubular mitochondrial AKT1 on metabolic syndrome-induced cardiomyopathy in transgenic mice. We generated transgenic mouse lines, KMioxCAKT, with inducible overexpression of mitochondrial AKT1, specifically in renal proximal tubular cells. One line was induced with tamoxifen, while the other was with corn oil. Both lines were fed a high-fat diet (HFD) for 16 weeks to provoke metabolic syndrome. For renal function, although serum BUN ( p= 0.728 ) and creatinine ( p=0.622 ) were not changed, the proteinuria ( p=0.031 ) and the urine KIM-1 ( p=0.012 ) were decreased in tamoxifen-induced KMioxCAKT. The glomerulosclerosis index ( p =0.027), tubulointerstitial fibrosis score ( p =0.032), tubular dilatation score ( p =0.032), tubular vacuolation score ( p =0.031), and tubular casts ( p =0.018) of renal histology were amended in tamoxifen-induced KMioxCAKT. The fasting glucose level ( p =0.015); the 15 to 120 minutes glucose levels ( p =0.020); the fasting insulin level ( p=0.037 ), and insulin resistance measured by Homeostatic Model Assessment for Insulin Resistance ( p=0.005 ) were reduced in metabolic syndrome with augmentation of renal tubular mitochondrial AKT1. In addition, tamoxifen-induced KMioxCAKT mice had significantly attenuated cardiac hypertrophy in the heart compared to corn oil-induced KMioxCAKT. Our results suggest that enhancing renal proximal tubular mitochondrial AKT1 could ameliorate the cardiorenal metabolic syndrome.
Abstract P2152: Asperuloside Ameliorates Obesity-associated Endothelial Dysfunction Via Activating Nrf2/HO-1 Signaling Pathway
Authors: Chufeng He
Journal: Circulation Research (2023)

Objectives: Asperuloside (ASP) is the major bioactive components in Eucommia species but its effects on vascular health and the underlying mechanism remain unknown. In this study we aim to investigate the protective vascular effect of ASP and we hypothesize that ASP ameliorates obesity associated endothelial dysfunction. Methods: After induction of obesity in C57BL/6 male mice fed with high fat diet for twelve weeks, they were treated daily with ASP (50 mg/kg) via oral gavage for seven weeks in vivo study. C57BL/6 mice aortae were used for ex vivo study and endothelial cells were used for in vitro study. Vascular function was determined by wire myograph. DHE and MitoSOX staining were used to detect ROS levels. QPCR, aortic en-face immunofluorescence staining and Western blotting were used to analyze mRNA and protein expressions. Small interfering (si)RNA of Nrf2 and HO-1, inhibitors of Nrf2 (ML385) and HO-1 (OB-24) were used to elucidate the mechanistic pathway of ASP ’s action. Findings: ASP significantly improved endothelial-dependent relaxations, suppressed oxidative stress, reduced VCAM-1 (~2 fold) and ICAM-1 (~2 fold) expression, increased both Nrf2 (~1.5 fold) and HO-1 (~1.5 fold) expression in obese mice aortae, IL-1β-treated aortae ex vivo and endothelial cells. However, ASP failed to prevent endothelial dysfunction in HO-1 inhibited aortae and HO-1-siRNA transfected endothelial cells, indicating that ASP improved endothelial function through upregulating HO-1. Nrf2 knockdown attenuated HO-1 upregulation by ASP, showing that ASP regulated Nrf2/HO-1 pathway. ASP increased nuclear Nrf2 and dual luciferase reporter assay illustrated that ASP increased ARE binding activity. Moreover, inhibition of Nrf2/ARE binding by ML385 blocked vascular protective effects and HO-1 upregulation by ASP, showing that ASP activated Nrf2/HO-1 signaling by stimulating Nrf2/ARE binding. Conclusions: This study for the first time shows that ASP rescues endothelial dysfunction and suppresses vascular oxidative stress associated with obesity and IL-1β via activating Nrf2/HO-1 pathway. These findings explore the pharmacological effects of ASP and contribute to the development of a novel treatment for endothelial dysfunction in cardiovascular diseases.
Abstract P2071: Mutation-Specific Calmodulin-Kinase II Activation In Hypertrophic Cardiomyopathy
Authors: Garrett T Hauck, Matthew Klass, Jil C Tardiff
Journal: Circulation Research (2023)

Autophosphorylation of Calmodulin-Kinase IIδ (CaMKIIδ) at Thr-287 leads to autoactivation such that kinase activity is autonomous of Ca 2+ /CaM. In the heart, CaMKIIδ functions to regulate Ca 2+ homeostasis by targeting Ca 2+ handling proteins, including phospholamban (PLB). In sarcomeric hypertrophic cardiomyopathy (HCM), disruption of Ca 2+ homeostasis and aberrant autoactivation of CaMKIIδ has been linked to disease progression. We have previously shown that the pathogenic role of autonomous CaMKIIδ in HCM is mutation-specific. Transgenic mice expressing the HCM mutation R92W in cardiac troponin T (cTnT) exhibited an increase in autonomous CaMKIIδ while cTnT-R92L mice did not. The potentially clinically relevant mechanism(s) underlying this mutation specificity are unresolved. We hypothesize that a potential trigger of autonomous CaMKIIδ is accelerated myofilament Ca 2+ dissociation kinetics. We have previously shown that cTnT-R92W accelerates Ca 2+ dissociation, while cTnT-R92L decelerates Ca 2+ dissociation. Here, we extended our study to include cTnT-I79N and cTnT-R94H to determine if Ca 2+ dissociation rates can be linked to autonomous CaMKIIδ activity and thus provide a potential novel target in HCM. Our data show that cTnT-I79N accelerates Ca 2+ dissociation, and cTnT-R94H decelerates Ca 2+ dissociation, similar to our earlier findings. To determine whether the mutation-specific changes in Ca 2+ dissociation alter CaMKIIδ activity, transgenic mice expressing cTnT-R94H and cTnT-I79N were probed for autophosphorylated CaMKIIδ and Thr-17 phosphorylated PLB. At 2 months, PLB phosphorylation is unchanged in cTnT-R94H and cTnT-I79N mice. At 4 months, cTnT-I79N mice exhibit a trending increase in autonomous CaMKIIδ and cTnT-R94H mice exhibit a modest increase, similar to cTnT-R92W positive controls. These trends are matched by PLB phosphorylation levels. At 6 months, cTnT-R94H mice exhibit normal autonomous CaMKIIδ levels. While future and ongoing work will explore changes in myocellular Ca 2+ homeostasis in more depth, these preliminary data suggest that, unlike cTnT-R94H mice, cTnT-I79N mice exhibit elevated autonomous CaMKIIδ and these mutation-specific differences may underlie observed phenotypic variation in patients.